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Microbiology

General Submission Requirements

Successful isolation of potential pathogens depends upon specimen selection and collection, proper transport, and timely delivery to the laboratory.  Whenever possible, specimens should be obtained before antibiotics or other antimicrobial agents have been administered. Please indicate the source of the specimen on the test to better assist in the diagnosis and treatment of the patient. Please consult Test Directory for additional instructions. 

Not all specimens contain clinically significant pathogens. Organism identification and antimicrobial susceptibility studies will be performed only on appropriate isolates at an additional charge. 

  • Specimen collection from normally sterile sites requires a needle puncture or a surgical procedure to decrease the chance of contamination. Do not submit syringes with needles attached. If a syringe must be submitted, remove needle, expel air, and recap syringe. Tape syringe plunger in position to prevent accidental movement. 
  • Because specimens are frequently and routinely collected from sites that are not sterile, the quality of culture results is directly dependent on the collection technique for urine, sputum, specimens from the nasopharynx and wounds.
  • Specimens from sites such as skin, mucous membranes and the gastrointestinal tract are populated by indigenous microflora. Microbiological tests will be directed at the isolation of specific pathogenic agents.

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Temperature Requirements

Appropriate storage and transport temperatures for clinical specimens are essential for successful isolation of organisms. Refer to the Quest Diagnostics Test Directory for transport temperatures for tests ordered. Note that Virology cultures, Mycoplasma/Ureaplasma cultures, and Helicobacter pylori culture require appropriate transport medium and storage at less than or equal to -70 degrees C if frozen. If a suitable freezer is not available, maintain these sample types refrigerated.

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Smear Collection

If both a smear and culture are ordered, perform collection procedures using two (2) swabs. Submit both swabs in appropriate culture transport medium.

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Bacteriology Swab and Vial Transport Systems

See the Infectious Disease Primary Care Specimen Collection Guide brochure available through a local Quest Diagnostics laboratory or representative.

  • We currently supply rayon-tipped swabs in transport systems containing Amies liquid (red cap) or Amies gel (blue cap) transport media, in plastic tubes. The swabs in gel transport systems (blue cap) may be used to culture for aerobic and/or anaerobic organisms. Liquid transport systems (red cap) are intended for use only with aerobic culture or antigen detection tests and should not be used for anaerobic culture. After collection, plunge the swab into the liquid or gel transport medium to prevent drying.
  • An “ESwab” (elution Swab) is the preferred collection device. This device features a flocked nylon tip that acts as a small brush. The ESwabs are simply snapped off into the screw-cap vial provided with multipurpose transport medium.
  • Tissue specimens and sterile body fluids should be submitted in Anaerobe Transport vials for aerobic and/or anaerobic cultures. 
  • Mini-tip swabs with a thin flexible wire shaft are used to collect nasopharyngeal and male urethral specimens. 
  • Tests for molecular detection of bacterial require special collection and transport systems.

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Transport of Infectious Substances

A.   Adherence to Regulatory Requirements

When sending a specimen suspected of containing a human, animal, or plant pathogen, please follow all current legal and regulatory requirements for interstate and air transportation for biohazardous material. The requirements include packaging the organism in a double-walled container.

B.   Labeling and Packaging

Indicate the organism suspected and relevant patient history on the requisition form. Known “Select Agents” must not be submitted to Quest Diagnostics.

C.   For Transportation Using Quest Diagnostics Logistics:

If Then
Routine bacterial isolates Submit in a screw-cap agar slant using Trypticase soy agar with or without 5% sheep blood or chocolate agar or in an appropriate bacterial transport swab system such as a blue-cap gel swab.
Fungal specimens Submit in a screw-cap agar slant of Sabouraud Dextrose agar.
Anaerobic organisms Submit in Chopped Meat Glucose or Thioglycollate Broth.
 
  • All tubes must be appropriately labeled and tightly capped. Do not submit isolates on petri dishes.
  • Use a separate bag for each isolate.
  • Each isolate must be placed in a Tape Seal 95kPa Specimen Transport Bag with the absorbent sheet pre-inserted in the bag.
  • Insert the specimen into the bag through the slit. Remove the adhesive backing from the tape seal area. Fold bag at slit and align lines so that they correspond.
  • To the seal bag, place the bag on a solid, flat surface. Press hard at the center of the tape seal area while maintaining this pressure, slide hands outward to the edges to seal.
  • Please use an “infectious substance” label on the bag.

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Infectious Substances and Select Agents

Under the 2006 rules most specimens for clinical testing may be classified as either (listed from low- to high-risk) Category A Infectious Substances, Category B Infectious Substances, or Exempt specimens. Category A Infectious Substances are infectious substance which are transported in a form that, when exposure to it occurs, is capable of causing permanent disability, life-threatening or fatal disease in otherwise healthy humans or animals. Culture isolates of Mycobacterium tuberculosis are an example of a Category A Infectious substance. See IATA3.6.2.2.2.1 and IATA Guidelines for more about Packing Instruction 650 and 49 CFR173.134 and 173.196.; Category B Infectious Substances are infectious substances which are not generally capable of causing permanent disability or life-threatening or fatal disease in otherwise healthy humans or animals when exposed. Category B Infectious Substances are Infectious Substances that are not considered to be Category A Infectious Substances. 

Those needing to transport infectious substances should check with the DOT, the U.S. Centers for Disease Control and Prevention (CDC) or public health authorities to determine classification of the specimen and, correspondingly, how the specimen should be packaged for transport. In addition, some air carriers may not consider certain specimens as suitable for air transport.

Select Agents

Certain biological agents have a special classification and are in the form of organisms, virus or toxins and are known to have a potential use as agents of bioterrorism as defined by the U.S. Department of Agriculture and Centers for Disease Control and Prevention. The most current information, including a list of select agents may be found at www.selectagents.gov. These agents require special notification and handling and must NOT be sent to Quest Diagnostics. A known select agent or toxin may only be sent or transferred to individuals or entities (such as certain medical laboratories) registered to possess, use, or transfer that agent or toxin. Clients must not submit known Agents to any Quest Diagnostics laboratory.

Most clinical samples (i.e., blood, wound, etc) from patients potentially infected with Select Agents can be collected and processed by routine Clinical Microbiology procedures. If environmental contamination with a CDC Select Agent is suspected, please contact local Public Health Department for more details on how to handle this sample type. Do not submit environmental samples potentially contaminated with Select Agents to Quest Diagnostics facilities. Specimens identified as select agents by Quest Diagnostics will not be returned.

Importing Specimens

Clients sending specimens from outside of the United States should contact the Quest Diagnostics laboratory before sending the specimen to the Quest laboratory. Among several requirements, importing specimens suspected of containing an infectious substance into the United States requires the receiving laboratory to have a valid CDC Import Permit and be labeled according to CDC requirements in addition to the DOT requirements discussed above. Importing specimens not suspected of containing a human pathogen requires a statement to that fact to accompany the specimen.

Specimens containing potential agents of bioterrorism as specified on the Commerce Control List will not be returned. The Commerce Control List is available from the Bureau of Industry and Security at www.bis.doc.gov.

Virology Swab and Vial Transport Systems

  1. We provide a multi-microbe specimen transport medium, V-C-M (Virus, Chlamydia, Mycoplasma), for the collection and transport of different types of specimens for viral isolation. (This medium will also support transport of Ureaplasma.) Store the V-C-M tube at room temperature until inoculated. Collection kits designed for specific anatomical sites are provided exclusively by Quest and are designed for genital, nasopharyngeal, and skin surface specimens.
  2. Do NOT use calcium alginate or wooden shaft swabs for specimen collection; use sterile rayon, Dacron® or flocked nylon swab on plastic or metal shafts.
  3. Write patient identification information and the source of the specimen on the Test Requisition and on the V-C-M (green-top) transport vial.
  4. Refrigerate specimens in V-C-M (green-top) tube immediately after collection.
  5. Keep specimens refrigerated for less than 48 hours, until the courier arrives. If a refrigerator is not available, keep specimen on wet ice or “cold packs” until it is picked up.
  6. If submission to the testing laboratory will take more than 48 hours, freeze the sample in V-C-M (green- top) tube at –70 °C, or colder, using dry ice. Transport it with enough dry ice to last 3 days. Freezing at –20 °C is not acceptable.
  7. Identify and separate virus isolation specimens from other specimens that are to be transported to the laboratory. Inform the courier as to the nature of specimens, so they may be appropriately transported to the laboratory. Transport refrigerated in a sterile leak-proof container. Do not freeze specimen unless in V-C-M transport medium.

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Parasitology & Stool Specimens, Storage and Transport

  1. While the ova and parasite (O&P exam) is ordered most often, it is not recommended as the routine test for diagnosis of intestinal parasites in the United States, as many intestinal parasites are not reliably detectable by O&P exam. An FAQ for the Diagnosis of Intestinal Parasites is available at Diagnosis of Intestinal Parasites | Quest Diagnostics 
  2. We provide Total-Fix® vials for the transport of stool for parasitology testing, including both antigen testing and traditional microscopy. Store and transport Total-Fix at room temperature.
  3. We provide Para-Pak® C&S vials for the transport of stool for cultures and antigen testing. Store and transport C&S refrigerated.
  4. For both transport media, transfer stool to the fill line on the transport vial as soon as possible and within two (2) hours of collection. 
  5. Transport urine in a sterile, leakproof container, refrigerated, for microscopy for Schistosoma.
  6. Transport sputum and bronchoalveolar lavages either unpreserved (refrigerated) or in 10% formalin (room temperature) for parasite microscopy.

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Mycology & Mycobacteriology Specimens, Storage and Transport

  1. For fungal cultures, submit specimens in a sterile, leakproof container, at room temperature. Do NOT refrigerate specimens.
  2. For mycobacterial (i.e., AFB) cultures, submit specimens in a sterile, leakproof container, refrigerated.
  3. If specimen is to be shared for mycobacterial and other cultures, split specimen, and transport portion for bacterial and/or fungal culture at room temperature. Transport specimen for mycobacterial culture refrigerated.
  4. For non–culture-based methodologies, refer to the Test Directory for transport instructions.

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Quick Reference: Microbiology Collection Instructions by Specimen Type

Specimen Type Instructions
Biopsy Specimens

Formalin-preserved or fixed-tissue specimens will not be accepted.

  • Collect fresh tissue using a set of separate sterile instruments. Each specimen need not be more than 1-2 cm in diameter.
  • Place each specimen into appropriate transport medium for the test(s) ordered.
  • If central nervous tissue is submitted, please inform the laboratory if the patient has or is suspected of having Creutzfeldt-Jacob Disease (CJD) by calling the laboratory and also indicating this on the Test Requisition.
Body Fluids Body fluids are collected only by a physician under aseptic conditions. Use extreme care in the collection of fluid. Do not contaminate, as any isolate from a sterile body fluid may be considered significant. Transfer fluid to appropriate transport medium for the test(s) ordered.
Cerebrospinal Fluid (CSF) CSF is collected only by a physician under aseptic conditions. After lumbar puncture, slowly drain the CSF fluid into the sterile leak-proof tubes. As a rule, the first tube is reserved for additional non-routine testing. The second tube or third tube should be sent to microbiology. Exception: Always send the most turbid tube to microbiology. Use extreme care in the collection of fluid. Do not contaminate, as any isolate from CSF may be considered significant. Viral testing of CSF should be performed by molecular testing. Transfer CSF to appropriate transport medium for the test(s) ordered.
Conjunctival/Ocular Collect samples using sterile rayon, Dacron® or flocked nylon swab with plastic or fine-gauge wire shaft. Do not use calcium alginate, cotton, or wooden shafted swabs which may inactivate the virus. Break, cut or fold the shaft to fit, if necessary.
Cutaneous or Vesicular Lesion

If a vesicle is present, disrupt the vesicle and collect the fluid with a swab. With the same swab, collect cells from the base of the vesicle by vigorous rubbing.

For non-vesicular lesions, remove scab or crusted material. Vigorously swab the base of the lesion to pick up infected cells.

Transfer the swab into appropriate transport medium for the test(s) ordered. Break, cut or fold the shaft to fit, if necessary.

Nasopharyngeal Swab Collect aspirates using a #5-8 disposable infant feeding tube attached to a 10 mL syringe or large suction bulb. If nasal material cannot be aspirated, instill up to 5 mL of saline into the nasal passages and re-aspirate to collect washings. As an alternative, use a suction catheter with a mucus trap. Place specimens in the appropriate transport medium for the test(s) ordered.
Rectal Swabs Insert swab 4–6 centimeters into rectum and roll swab against the mucosa. Examine the swab to help ensure that fecal material is not present. Place swab in appropriate transport medium for the test(s) ordered.
Respiratory Aspirate or Washings Collect aspirates using a #5–8 disposable infant feeding tube attached to a 10 mL syringe or large suction bulb. If nasal material cannot be aspirated, instill up to 5 mL of saline into the nasal passages and re-aspirate to collect washings. As an alternative, use a suction catheter with a mucus trap. Place specimens in the appropriate transport medium for the test(s) ordered.
Stool Collect specimen using a stool "hat." Transfer stool to the fill line of the appropriate transport medium for the test(s) ordered within 30 minutes of collection.
Throat Swab Vigorously swab tonsillar area and posterior oropharynx using a sterile rayon, Dacron® or flocked nylon swab. Place swab in appropriate transport medium for the test(s) ordered.
Urethral Swab Insert flocked nylon or moistened mini-tipped swab at least 2 cm into urethral orifice. Rotate gently to obtain epithelial cells. Place swab into appropriate transport medium for the test(s) ordered.
Urine Obtain a fresh, clean-catch specimen in a clean container. If centrifugation is required, transfer about 10 mL to a centrifuge tube and centrifuge for at least 15 minutes in a centrifuge supplied by Quest, which can develop a relative centrifugal force equivalent to between 1450 x g (fixed-angle rotor) and 1600 x g (horizontal rotor) when operating within the instrument manufacturer’s specifications. Transfer specimen to the appropriate transport medium for the test(s) ordered. If centrifuged, transfer about 2 mL, or as much of the sediment as possible. Discard the remainder.

Quick Reference: Popular Microbiology Collection Containers